Crystal structures of histone demethylase JMJD2A reveal basis for substrate specificity


Post-translational histone modification has a fundamental role in chromatin biology and is proposed to constitute a ‘histone code’ in epigenetic regulation1,2. Differential methylation of histone H3 and H4 lysyl residues regulates processes including heterochromatin formation, X-chromosome inactivation, genome imprinting, DNA repair and transcriptional regulation3. The discovery of lysyl demethylases using flavin (amine oxidases)4 or Fe(ii) and 2-oxoglutarate as cofactors (2OG oxygenases)5,6,7 has changed the view of methylation as a stable epigenetic marker. However, little is known about how the demethylases are selective for particular lysyl-containing sequences in specific methylation states, a key to understanding their functions. Here we reveal how human JMJD2A (jumonji domain containing 2A), which is selective towards tri- and dimethylated histone H3 lysyl residues 9 and 36 (H3K9me3/me2 and H3K36me3/me2), discriminates between methylation states and achieves sequence selectivity for H3K9. We report structures of JMJD2A–Ni(ii)–Zn(ii) inhibitor complexes bound to tri-, di- and monomethyl forms of H3K9 and the trimethyl form of H3K36. The structures reveal a lysyl-binding pocket in which substrates are bound in distinct bent conformations involving the Zn-binding site. We propose a mechanism for achieving methylation state selectivity involving the orientation of the substrate methyl groups towards a ferryl intermediate. The results suggest distinct recognition mechanisms in different demethylase subfamilies and provide a starting point to develop chemical tools for drug discovery and to study and dissect the complexity of reversible histone methylation and its role in chromatin biology.

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Figure 1: Overview of the H3K9 peptide bound to the catalytic domain of JMJD2A.
Figure 2: JMJD2A substrate peptide binding.
Figure 3: Mass spectra (MALDI–TOF) of JMJD2A demethylation reactions with native and modified histone peptides.
Figure 4: Active site and mechanism of JMJD2A.


  1. 1

    Margueron, R., Trojer, P. & Reinberg, D. The key to development: interpreting the histone code? Curr. Opin. Genet. Dev. 15, 163–176 (2005)

  2. 2

    Zhang, Y. & Reinberg, D. Transcription regulation by histone methylation: interplay between different covalent modifications of the core histone tails. Genes Dev. 15, 2343–2360 (2001)

  3. 3

    Martin, C. & Zhang, Y. The diverse functions of histone lysine methylation. Nature Rev. Mol. Cell Biol. 6, 838–849 (2005)

  4. 4

    Shi, Y. et al. Histone demethylation mediated by the nuclear amine oxidase homolog LSD1. Cell 119, 941–953 (2004)

  5. 5

    Tsukada, Y. et al. Histone demethylation by a family of JmjC domain-containing proteins. Nature 439, 811–816 (2006)

  6. 6

    Klose, R. J., Kallin, E. M. & Zhang, Y. JmjC-domain-containing proteins and histone demethylation. Nature Rev. Genet. 7, 715–727 (2006)

  7. 7

    Whetstine, J. R. et al. Reversal of histone lysine trimethylation by the JMJD2 family of histone demethylases. Cell 125, 467–481 (2006)

  8. 8

    Flashman, E. & Schofield, C. J. The most versatile of all reactive intermediates? Nature Chem. Biol. 3, 86–87 (2007)

  9. 9

    Chen, Z. et al. Structural insights into histone demethylation by JMJD2 family members. Cell 125, 691–702 (2006)

  10. 10

    Clifton, I. J. et al. Structural studies on 2-oxoglutarate oxygenases and related double-stranded β-helix fold proteins. J. Inorg. Biochem. 100, 644–669 (2006)

  11. 11

    Lo, W. S. et al. Snf1–a histone kinase that works in concert with the histone acetyltransferase Gcn5 to regulate transcription. Science 293, 1142–1146 (2001)

  12. 12

    Nelson, C. J., Santos-Rosa, H. & Kouzarides, T. Proline isomerization of histone H3 regulates lysine methylation and gene expression. Cell 126, 905–916 (2006)

  13. 13

    Price, J. C., Barr, E. W., Tirupati, B., Bollinger, J. M. & Krebs, C. Kinetic dissection of the catalytic mechanism of taurine:α-ketoglutarate dioxygenase (TauD) from Escherichia coli. Biochemistry 42, 7497–7508 (2003)

  14. 14

    Grzyska, P. K. et al. Steady-state and transient kinetic analyses of taurine/α-ketoglutarate dioxygenase: effects of oxygen concentration, alternative sulfonates, and active-site variants on the FeIV-oxo intermediate. Biochemistry 44, 3845–3855 (2005)

  15. 15

    Elkins, J. M. et al. Structure of Factor-inhibiting Hypoxia-inducible Factor (HIF) reveals mechanism of oxidative modification of HIF-1. J. Biol. Chem. 278, 1802–1806 (2003)

  16. 16

    Liang, G., Klose, R. J., Gardner, K. E. & Zhang, Y. Yeast Jhd2p is a histone H3 Lys4 trimethyl demethylase. Nature Struct. Mol. Biol. 14, 243–245 (2007)

  17. 17

    Seward, D. J. et al. Demethylation of trimethylated histone H3 Lys4 in vivo by JARID1 JmjC proteins. Nature Struct. Mol. Biol. 14, 240–242 (2007)

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We thank A. Edwards and J. Min for critical discussions and reading of the manuscript; P. Siegbahn for support; and F. Sobott, K. Di Gleria and C. Webby for technical help with MALDI–TOF analysis. The Structural Genomics Consortium is a registered charity funded by the Wellcome Trust, GlaxoSmithKline, Genome Canada, the Canadian Institutes of Health Research, the Ontario Innovation Trust, the Ontario Research and Development Challenge Fund, the Canadian Foundation for Innovation, Vinnova, Knut and Alice Wallenberg foundation and Karolinska Institutet. C.J.S. and co-workers are funded by the Wellcome Trust, and the Biotechnology and Biological Sciences Research Council, United Kingdom. Atomic coordinates and structure factors for the reported crystal structures have been deposited with the Protein Data Bank under accession codes 2oq7 (JMJD2A–NOG–Ni(II)), 2oq6 (JMJD2A–NOG–Ni(II)–H3K9me3), 2os2(JMJD2A–NOG–Ni(II)–H3K36me3), 2ot7(JMJD2A–NOG–Ni(II)–H3K9me1) and 2ox0 (JMJD2A–NOG–Ni(II)–H3K9me2).

Author Contributions S.S.N. purified, crystallized, collected data and performed MALDI–TOF analyses; K.L.K., M.A.M. and E.S.P. collected, processed and refined X-ray data; D.B. and B.M.R.L. performed MS analysis; P.S. and O.G. cloned the construct; F.v.D. collected data; N.R.R., J.C.S. and J.O. synthesized peptides; T.B. performed MD studies; J.E.B. performed bioinformatic analyses; M.S. was involved in study design; C.J.S. and U.O. designed the study, analysed data and wrote the paper. M.A.M. and D.B. contributed equally to the study. All authors discussed the results and commented on the manuscript.

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Correspondence to Christopher J. Schofield or Udo Oppermann.

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C.J.S. is a co-founder and director of ReOx, a company that aims to exploit knowledge about the hypoxic response for therapeutic benefit.

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This file contains Supplementary Tables S1-S2 and Supplementary Figures S1-S10 with Legends. (PDF 1841 kb)

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Ng, S., Kavanagh, K., McDonough, M. et al. Crystal structures of histone demethylase JMJD2A reveal basis for substrate specificity. Nature 448, 87–91 (2007).

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